The RNA-binding protein hnRNPLL induces a T cell alternative splicing program delineated by differential intron retention in polyadenylated RNA

Abstract

BackgroundRetention of a subset of introns in spliced polyadenylated mRNA is emerging as a frequent, unexplained finding from RNA deep sequencing in mammalian cells.ResultsHere we analyze intron retention in T lymphocytes by deep sequencing polyadenylated RNA. We show a developmentally regulated RNA-binding protein, hnRNPLL, induces retention of specific introns by sequencing RNA from T cells with an inactivating Hnrpll mutation and from B lymphocytes that physiologically downregulate Hnrpll during their differentiation. In Ptprc mRNA encoding the tyrosine phosphatase CD45, hnRNPLL induces selective retention of introns flanking exons 4 to 6; these correspond to the cassette exons containing hnRNPLL binding sites that are skipped in cells with normal, but not mutant or low, hnRNPLL. We identify similar patterns of hnRNPLL-induced differential intron retention flanking alternative exons in 14 other genes, representing novel elements of the hnRNPLL-induced splicing program in T cells. Retroviral expression of a normally spliced cDNA for one of these targets, Senp2, partially corrects the survival defect of Hnrpll-mutant T cells. We find that integrating a number of computational methods to detect genes with differentially retained introns provides a strategy to enrich for alternatively spliced exons in mammalian RNA-seq data, when complemented by RNA-seq analysis of purified cells with experimentally perturbed RNA-binding proteins.ConclusionsOur findings demonstrate that intron retention in mRNA is induced by specific RNA-binding proteins and suggest a biological significance for this process in marking exons that are poised for alternative splicing.

Global analysis of physical and functional RNA targets of hnRNP L reveals distinct sequence and epigenetic features of repressed and enhanced exons

“…Furthermore, hnRNP LL, a paralog of hnRNP L, has been shown to regulate at least some intron retention events in primary mouse T cells (Cho et al 2014). However, our rMATS analysis only predicted ∼100 intron retention events impacted by hnRNP L, many fewer than the number of cassette exons that are regulated (Table 1).…”

The RNA-binding profile of Acinus, a peripheral component of the exon junction complex, reveals its role in splicing regulation

“…Intron retention also plays a role in cancer as a mechanism of tumor suppressor inactivation (Jung et al 2015). Since Acinus controls the splicing of a subset of introns, it could represent a good candidate for the specific control of IR in a precise biological context, as was observed for hnRNPLL in intron retention regulation in T cells (Cho et al 2014). However, contrary to the known tissue-specific expression of hnRNPLL, the ACIN1 gene appears to be ubiquitously expressed (data not shown).…”

Intron retention in mRNA: No longer nonsense

“…From studies in neurons , T cells , and our own data in granulocytes , the retention of introns in mRNA transcripts is evidently a regulated process with possibly diverse functions in normal cells. As discussed above, intron‐retaining transcripts can play a role in the regulation of gene expression or produce novel protein isoforms .…”

“…IR is the least described mode of splicing in humans and animals; however, its role has been studied more extensively in recent years. Although IR occurs less frequently than other modes of AS, it serves essential functions in the regulation of dosage compensation of the X chromosome , tissue specific protein diversity , mRNA localization , splicing regulation , and gene expression control (Table ).…”